The present invention relates generally to the field of medicine, and relates specifically to angiogenesis and metastasis of cancer tissues. Specifically the invention relates to the use of antagonists of the serine integral membrane protease, dipeptidyl peptidase IV (DPPIV also known as CD26).
Growth of new blood vessels (angiogenesis) plays a key role in tissue repair and in cancer progression. The invasion of cells into a connective tissue barrier during angiogenesis requires remodeling of the extracellular matrix (ECM) by migratory cells (Martin, 1997). In cancer invasion such cellular activities occur on membrane protrusions invadopodia (Chen, 1979) which exhibit dynamic membrane mobility, ECM adhesion and degradation. Thus, cellular invasion is an important process for cancer metastasis (Stetler-Stevenson et al., 1993). Several classes of proteases including matrix metalloproteinases (MMPs), serine proteases, cysteine proteases (cathepsin B and cathepsin L), and aspartic acid proteases (cathepsin D) can degrade proteins in the ECM (Chen, 1992). And invading cancer cells possess ECM degrading proteolytic enzymes that are concentrated at specialized plasma membrane protrusions, termed invadopodia (Chen et al., 1994). Recent studies showed that integral membrane proteases might contribute significantly to ECM degradation and ultimately cancer invasion by virtue of their location at invadopodia (Monsky and Chen, 1993).
Recent evidence has demonstrated the involvement of serine-integral membrane roteases (SIMP), including dipeptidyl peptidase IV (DPPIV)/CD26 and seprase, in cell surface proteolysis (Chen, 1996). SIMP members are type II transmembrane proteins, with cytoplasmic tails that contain 6 amino acids (a.a.) followed by a 20 a.a. (seprase) or 22 a.a. (DPPIV) transmembrane domain at the N-terminus and a stretch of 200 a.a. at the C-terminus that constitutes a catalytic region with the catalytic serine in a non-classical orientation (Goldstein et al., 1997; Pineiro-Sanchez et al., 1997).
DPPIV specifically removes N-terminal dipeptides from oligo-peptides, which include Neuro-Peptide Y and other peptide hormones, with either L-proline, L-hydroxyproline, or L-alanine at the penultimate position (Heins et al., 1988, Walter et al., 1980). DPPIV has been shown to be an adhesion receptor for collagen (Bauvois, 1988; Hanski et al., 1988; Loster et al., 1995) or fibronectin (Cheng et al., 1998; Johnson, et al., 1993; Piazza et al., 1989). In addition, a recent report showed that DPPIV also possesses a seprase-like gelatinase activity and therefore endopeptidase activity (Bermpohl et al., 1998), suggesting its involvement in collagen degradation. DPPIV is expressed constitutively on brush border membranes of intestine and kidney epithelial cells (Yaron and Naider, 1993; Morimoto and Schlossman, 1994).
Seprase, originally identified as a 170 kDa membrane-bound gelatinase is expressed on invadopodia of highly aggressive melanoma LOX cells (Aoyama and Chen, 1990; Mueller et al., 1999; Monsky et al., 1994). The active enzyme is a homodimer of 97 kDa subunits, which are proteolytically inactive (Pineiro-Sanchez et al., 1997). Analysis of the deduced amino acid sequence from a cDNA that encodes the 97 kDa subunit (Goldstein et al., 1997) revealed that it is homologous to DPPIV, and is essentially identical to fibroblast activation protein xcex1 (FAPxcex1) (Scanlan et al., 1994), which is expressed on reactive stromal fibroblasts of epithelial cancers and healing wounds (Garin-Chesa et al., 1990). In addition, DNA and protein analysis of embryonic tissues has suggested potential additional members of SIMP (Bermpohl et al., 1998).
A growing body of evidence indicates that angiogenesis is essential to the progression of cancer. Angiogenesis is the sprouting of new capillaries from preexisting blood vessels. Normally, angiogenesis in mammals is confined to the reproductive system, embryogenesis and development, and repair after injury. However, angiogenesis can also occur in pathological conditions such as cancer, retinal neovascularization, neovascularization in atherosclerotic plaques, hemangiomas, arthritis, and psoriasis. See Folkman, 1995. Without vascularization, tumors may remain for years as small (less than a few millimeters) asymptomatic lesions. Weidner et al. (1991). Angiogenesis allows the cancer cells access to the circulatory system. The new blood vessels provide a gateway for cancer cells to enter the circulation and metastasize to distant sites (Folkman 1990; Klagsbrunn and Soker, 1993).
As in cancer cell invasion, angiogenesis involves matrix degradation by migrating endothelial cells at the invasion front; proteases including matrix metalloproteases (MMPs) (Hiraoka et al., 1998; Brooks et al., 1998) and plasminogen activators (Pepper et al., 1993) are essential but novel membrane-bound proteases active at sites of angiogenesis are yet to be defined.
Several approaches for inhibition of angiogenesis have been proposed as useful therapies for restricting tumor growth. These include inhibition of angiogenesis by (1) inhibition of release of xe2x80x9cangiogenic moleculesxe2x80x9d such as VEGF (Vascular endothelial growth factor) and basic.FGF (fibroblast growth factor), (2) neutralization of angiogenic molecules, such as by use of anti-b.FGF antibodies, (3) targeted inhibition on .alpha..sub.v .beta..sub.3 integrin, and (4) inhibition of the endothelial cell response to angiogenic stimuli. This latter strategy has received attention, and Folkman et al., Cancer Biology, 3:89-96 (1992), have described several endothelial cell response inhibitors, including collagenase inhibitor, basement membrane turnover inhibitors, angiostatic steroids, fungal-derived angiogenesis inhibitors, platelet factor 4, thrombospondin, arthritis drugs such as D-penicillamine and gold thiomalate, vitamin D.sub.3 analogs, alpha-interferon, and the like that might be used to inhibit angiogenesis.
Monoclonal antibodies (MAbs) to human tumor-associated differentiation antigens offer promises for the xe2x80x9ctargetingxe2x80x9d of various antitumor agents such as radioisotopes, chemotherapeutic drugs, and toxins. [Order, in xe2x80x9cMonoclonal Antibodies for Cancer Detection and Therapyxe2x80x9d, Baldwin and Byers, (eds.),London, Academic Press (1985)].
In addition, some monoclonal antibodies have the advantage of killing tumor cells via antibody-dependent cellular cytotoxicity (ADCC) or complement-dependent cytotoxicity (CDC) in the presence of human effector cells or serum [Hellstrom et al., Proc. Natl. Acad. Sci. USA 83:7059-7063 (1986)], and there are a few monoclonal antibodies that have a direct antitumor activity which does not depend on any host component [Drebin et al., Oncogene 2:387-394 (1988)].
For additional proposed inhibitors of angiogenesis, see Blood et al., Bioch. Biophys. Acta., 1032:89-118 (1990) for a general review of angiogenesis and tumor metastasis; also Moses et al., Science, 248:1408-1410 (1990) describes a protein inhibitor of angiogenesis derived from cartilage; and Ingber and Folkman, Lab. Invest. 59:44-51 (1988) describes inhibition of angiogenesis through modulation of collagen metabolism.
U.S. Pat. No. 5,092,885, of Yamada et al. discloses laminin peptides with angiogenesis-blocking activity. U.S. Pat. No. 5,112,946 of Maione et al. discloses modified PF4 compositions as inhibitors of angiogenesis.
U.S. Pat. No. 5,192,744, discloses human thrombospondin for use as an inhibitor of angiogenesis. U.S. Pat. No. 5,202,352 discloses intravascular embolizing agents containing angiogenesis inhibiting substances in oils, emulsions or suspensions. U.S. Pat. No. 5,766,591 discloses antagonists of vitronectin alpha.sub.v.beta.sub.3 as angiogenesis inhibitors.
U.S. Pat. No. 5,980,896 of Hellstrom et al. discloses antibodies and immunoconjugates reactive with human carcinomas and is especially useful in practicing the full scope of the present invention. Among the disclosed compositions and methods which are especially applicable to the present invention are: chimeric antibodies, immunoconjugates thereof and their methods of preparation and use; and anti-tumor drugs, cytotoxins, radioactive agents and enzymes useful in immunoconjugate compositions. The text of U.S. Pat. No. 5,980,896 is hereby incorporated by reference in its entirety.
There is still a need, however for novel and more effective anti-angiogenesis therapies for use alone or in combination with one or more of the currently available therapies for treatment of growth and proliferative disorders involving angiogenesis.
The invention provides monospecific antibodies which specifically bind an epitope of a human DPPIV (dipeptidyl peptidase IV/CD26) and inhibits angiogenesis.
Also provided are bispecific antibodies with binding specificity for two epitopes, one of which is an epitope of DPPIV. The bispecific antibodies of the present invention include those in which the second epitope bound is an epitope of seprase, MT1-MMP, MMP-2 or xcex1(3)xcex2(1)-integrin.
The present invention further provides immunoconjugates comprising a monospecific or a bispecific antibody which specifically binds an epitope of human DPPIV (dipeptidyl peptidase IV/CD26) and inhibits angiogenesis, joined to a therapeutic agent.
In yet another aspect the invention provides pharmaceutical compositions for inhibiting angiogenesis comprising an effective amount of an antibody which specifically binds an. epitope of a human DPPIV (dipeptidyl peptidase IV/CD26) and inhibits angiogenesis, the antibody being formulated in a pharmaceutically acceptable carrier.
In yet another aspect the invention provides pharmaceutical compositions for inhibiting angiogenesis comprising an effective amount of an immunoconjugate of an antibody which specifically binds an epitope of a human DPPIV (dipeptidyl peptidase IV/CD26) and inhibits angiogenesis, the antibody being formulated in a pharmaceutically acceptable carrier.
The present invention yet further provides a method of treating a patient suffering from a growth or proliferative disorder involving angiogenesis, comprising administering to the site of angiogenesis an effective amount of an antibody which specifically binds an epitope of a human DPPIV (dipeptidyl peptidase IV/CD26) and inhibits angiogenesis.
In yet another aspect the present invention provides a method of treating a patient suffering from a growth or proliferative disorder involving angiogenesis, comprising administering to the site of angiogenesis an effective amount of an immunoconjugate which specifically binds an epitope of a human DPPIV (dipeptidyl peptidase IV/CD26) and inhibits angiogenesis.
In a further aspect the present invention provides continuous cell lines which produce monospecific antibodies that specifically bind an epitope of a human DPPIV (dipeptidyl peptidase IV/CD26) and thereby inhibit angiogenesis.